Evaluation of plasma exosomal microRNA-21 as a circulating biomarker in oesophageal adenocarcinoma (#54)

Oesophageal adenocarcinoma generally has a poor prognosis, although some patients benefit from chemotherapy prior to surgery. MicroRNAs are associated with tumorigenesis, metastasis and chemotherapy resistance. For some cancer types, circulating tumor-derived microRNAs encapsulated in exosomes have been reported in plasma and serum. These microRNA profiles might predict which oesophageal adenocarcinoma patients will respond to chemotherapy, but there are no reports of tumour-derived circulating microRNAs in oesophageal cancer. We tested methods for isolating exosomes from plasma and serum, and we compared the level of oesophageal adenocarcinoma-associated microRNA-21 in plasma exosomes from adenocarcinoma patients with those from healthy controls.

Plasma and serum samples were centrifuged at 16,000 g to remove cellular debris. The supernatant was used for ultracentrifugation or Exoquick™ protocols to pellet exosomes. Particle count and size were determined by NanoSight Nanoparticle Tracking Analysis. To normalise microRNA levels between samples, a synthetic microRNA was spiked in prior to RNA extraction. MicroRNA levels were measured using relative quantitative reverse-transcriptase polymerase chain reaction.

Exoquick pellets had significantly more particles (average 1130 vs 29 particles), with significantly smaller size compared with ultracentrifugation pellets (p<0.05). MicroRNA-21 levels were average 7 times higher in Exoquick™ pellets than in ultracentrifugation pellets. MicroRNA-21 levels in Exoquick™ pellets from plasma were average 4.5 times higher than in serum. There was no significant difference in plasma exosomal microRNA-21 levels between 10 healthy controls and 10 oesophageal adenocarcinoma patients (p=0.79). However, the miR-21 level was significantly correlated with the particle concentration in the exosomal pellets (rho=0.79, p<0.0001).

Exoquick™ may be preferable to ultracentrifugation because it yields more particles with a size consistent with exosomes. The level of cancer-associated microRNA-21 in plasma circulating exosomes is not a suitable biomarker for oesophageal adenocarcinoma.